Two aspects of the relationship between chromosome structure and composition and chromosome function are under investigation. The first is analysis of changes in chromosome structure which accompany chromosome functions such as replication, transcription and chromatin folding. In order to approach this problem we are developing techniques to localize at the electron microscopic level specific chromatin-associated proteins and specific nucleic acid sequences in condensed and dispersed chromatin. Our test system uses antibody against histone H1 and its interaction with chromatin as assayed by electron microscopic visualization of ferritin-conjugated antibody-antigen complexes. We are optimizing conditions for such reactions in order to localize and quantitate H1-chromatin interactions under various conditions. We are extending standard in situ hybridization technology in order to allow localization of both DNA and nascent RNA sequences of eukaryotic chromosomes. The test system for this work involves in situ hybridization of complementary RNA made from mouse satellite DNA to metaphase chromosomes immobilized on an electron microscope grid. The satellite is known to be located at the centromere region but its precise location is unknown. Other studies in the laboratory on the DNA sequences and non-histone chromosomal proteins restricted to the centromere region correlate with the in situ experiment since they are directed toward the elucidation of putative DNA sequence-specific protein interactions in the centromere which may be involved in chromosome folding, placement of structures required for chromosome segregation and/or chromosome pairing.